Minimally invasive enteric pathogens interact with the intestinal tract at the epithelium but do not invade deeper mucosal layers or other organs. Since they have little contact with phagocytes, which are key for detecting and eliminating invasive pathogens, we hypothesize that the interactions of minimally invasive pathogens with the intestinal epithelium are central for initiating host immune responses and producing and/or delivering antimicrobial effectors. The overall goal of our studies is to define the importance and mechanisms of these interactions, and to apply such insights to the prevention and treatment of diseases caused by these pathogens. The proposed studies will focus on Giardia as a leading protozoan cause of diarrheal disease worldwide and a model of a non-invasive microbe that causes little mucosal inflammation, and Cryptosporidium parvum, which is a major cause of waterborne diarrheal disease, and invades the intestinal epithelium, but not deeper mucosal layers, and causes mucosal inflammation. The proposed studies build on our findings in the prior Program period and are a direct continuation of that work. The studies have the following Specific Aims: Aim 1. To identify new Giardia gene products targeted by antigiardial IgA and determine their importance in eliciting antigiardial immunity. Based on our prior finding that IgA is central for antigiardial host defense, these studies will use a new mouse model of hyperimmune serum IgA production and 2-D gels and mass spectrometry to identify new giardial antigens, and test their importance for eliciting immune protection. Aim 2. To define the function of histamine in host defense against Giardia and identify new genes involved in antigiardial immune defense. Preliminary microarray data, together with previous reports, indicate that mast cells are important in antigiardial host defense. Based on this, we will test the hypothesis that the major mast cell product, histamine, is a key regulator of immune defense against Giardia. Aim 3. To define the importance of epithelial cell NF-kappaB in cryptosporidiosis and the mechanisms of IFN-gamma function in host defense against C. parvum. We have generated a new conditional knock-out mouse model of impaired NF-kappaB activation in intestinal epithelial cells, and will use it to test the hypothesis that NF-kappaB in these cells has important functions in host defense against C. parvum. Other studies will define mechanisms by which IFN-gamma, a key regulator of C. parvum host defense, exerts its functions.